Transcriptome analysis implicates involvement of long noncoding RNAs in cytoplasmic male sterility and fertility restoration in cotton

Abstract Background Hybrid cotton has greatly contributed to global increase in cotton productivity. The cytoplasmic male sterility (CMS)/restorer-of-fertility system is an important tool to exploit heterosis because it is convenient for commercial hybrid seed production. The importance of long noncoding RNAs (lncRNAs) in plant development is recognized, few analyses of lncRNAs during anther development of three-line hybrid cotton (CMS-D2 line A, maintainer line B, restorer-of-fertility line R) have been reported. Results Here, we performed transcriptome sequencing during anther development in Upland cotton carrying cytoplasmic male sterile Gossypium harknessii (D2) cytoplasm. Totally 80,695 lncRNAs were identified, in which 43,347 and 44,739 lncRNAs were differentially expressed in A–B and A–R comparisons, respectively. These lncRNAs represent functional candidates involved in CMS and fertility restoration. Gene ontology enrichment analysis indicated that cellular hormone metabolic processes and oxidation–reduction reaction processes might be involved in CMS, and cellular component morphogenesis and small molecular biosynthetic processes might participate in fertility restoration. Analysis of the putative relationship between lncRNAs and miRNAs revealed that 63 lncRNAs were identified as putative precursors of 35 miRNAs, and qRT-PCR analysis showed a similar expression pattern to that of RNA-sequencing data. Furthermore, construction of lncRNA regulatory networks indicated that several miRNA–lncRNA–mRNA networks might be involved in CMS and fertility restoration. Conclusion Our findings provide systematic identification of lncRNAs during anther development and lays a solid foundation for future investigation of the regulatory molecular mechanisms and utilization in breeding of hybrid cotton.

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Transcriptome Analysis Implicates Involvement of Long Noncoding RNAs in Cytoplasmic Male Sterility and Fertility Restoration in Cotton

International Journal of Molecular Sciences ◽

10.3390/ijms20225530 ◽

2019 ◽

Vol 20 (22) ◽

pp. 5530

Author(s):

Bingbing Zhang ◽

Xuexian Zhang ◽

Meng Zhang ◽

Liping Guo ◽

Tingxiang Qi ◽

...

Keyword(s):

Cytoplasmic Male Sterility ◽

Male Sterility ◽

Regulatory Networks ◽

Fertility Restoration ◽

Noncoding Rnas ◽

Reduction Reaction ◽

Anther Development ◽

Long Noncoding Rnas ◽

Hybrid Seed Production ◽

Restorer Of Fertility

The cytoplasmic male sterility (CMS)/restorer-of-fertility system is an important tool to exploit heterosis during commercially hybrid seed production. The importance of long noncoding RNAs (lncRNAs) in plant development is recognized, but few analyses of lncRNAs during anther development of three-line hybrid cotton (CMS-D2 line A, maintainer line B, restorer-of-fertility line R) have been reported. Here, we performed transcriptome sequencing during anther development in three-line hybrid cotton. A total of 80,695 lncRNAs were identified, in which 43,347 and 44,739 lncRNAs were differentially expressed in A–B and A–R comparisons, respectively. These lncRNAs represent functional candidates involved in CMS and fertility restoration. GO analysis indicated that cellular hormone metabolic processes and oxidation–reduction reaction processes might be involved in CMS, and cellular component morphogenesis and small molecular biosynthetic processes might participate in fertility restoration. Additionally, 63 lncRNAs were identified as putative precursors of 35 miRNAs, and quantitative reverse transcription polymerase chain reaction (qRT-PCR) showed a similar expression pattern to RNA-seq data. Furthermore, construction of lncRNA regulatory networks indicated that several miRNA–lncRNA–mRNA networks might be involved in CMS and fertility restoration. Our findings provide systematic identification of lncRNAs during anther development and lays a solid foundation for the regulatory mechanisms and utilization in hybrid cotton breeding.

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Selection of Nuclear Genotypes Associated with the Thermo-inducibility of Owen-type Cytoplasmic Male Sterility in Sugar Beet (Beta vulgaris L.)

10.21203/rs.3.rs-551730/v1 ◽

2021 ◽

Author(s):

Hiroaki Matsuhira ◽

Kazuyoshi Kitazaki ◽

Katsunori Matsui ◽

Keisi Kubota ◽

Yosuke Kuroda ◽

...

Keyword(s):

Sugar Beet ◽

Cytoplasmic Male Sterility ◽

Male Sterility ◽

Beta Vulgaris ◽

Parental Line ◽

Hybrid Seed Production ◽

Male Sterile ◽

Flowering Season ◽

Restorer Of Fertility ◽

The Stability

Abstract The stability of cytoplasmic male sterility expression in several genetic backgrounds was investigated in sugar beet (Beta vulgaris L.). Nine genetically heterogenous plants from old cultivars were crossed with a cytoplasmic male-sterile line to obtain 266 F1 plants. Based on marker analysis using a multiallelic DNA marker linked to restorer-of-fertility 1 (Rf1), we divided the F1 plants into 15 genotypes. We evaluated the phenotypes of the F1 plants under two environmental conditions: greenhouse rooms with or without daytime heating during the flowering season. Three phenotypic groups appeared: those consistently expressing male sterility (MS), those consistently having restored pollen fertility, and those expressing MS in a thermo-sensitive manner. All plants in the consistently male-sterile group inherited a specific Rf1 marker type named p4. We tested the potential for thermo-induced male-sterile plants to serve as seed parents for hybrid seed production, and three genotypes were selected. Open pollination by a pollen parental line with a dominant trait of red-pigmented hypocotyls and leaf veins resulted in seed setting on thermo-induced male-sterile plants, indicating that their female organs were functional. More than 99.9% of the progeny expressed the red pigmentation trait; hence, highly pure hybrids were obtained. We determined the nucleotide sequences of Rf1 from the three genotypes: one had a novel allele and two had known alleles, of which one was reported to have been selected previously as a nonrestoring allele at a single US breeding station but not at other stations in the US, or in Europe or Japan, suggesting environmental sensitivity.

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Alloplasmic male sterile Brassica napus with Enarthrocarpus lyratus cytoplasm: introgression and molecular mapping of an E. lyratus chromosome segment carrying a fertility restoring gene

Genome ◽

10.1139/g03-055 ◽

2003 ◽

Vol 46 (5) ◽

pp. 792-797 ◽

Cited By ~ 15

Author(s):

H S Janeja ◽

S K Banga ◽

P B Bhaskar ◽

S S Banga

Keyword(s):

Brassica Napus ◽

Cytoplasmic Male Sterility ◽

Male Sterility ◽

Molecular Mapping ◽

Bulked Segregant Analysis ◽

Fertility Restoration ◽

Hybrid Seed Production ◽

Male Sterile ◽

Fertility Restorer ◽

Donor Species

A cytoplasmic male sterility (CMS) system for Brassica napus (2n = 38; AACC) was developed by backcross substitution of its nucleus into the cytoplasm of a wild crucifer, Enarthrocarpus lyratus. Male sterility was complete, stable, and expressed in small flowers with rudimentary anthers. Since the B. napus germplasm lines were complete or partial maintainers of male sterility, the required fertility restorer gene (Rfl) was introgressed from the cytoplasm donor species. Inheritance studies carried out on F1 and F2 populations derived from hybridizing cytoplasmic male sterile and male fertile near-isogenic (PNILs) lines of B. napus 'Westar', revealed a monogenic dominant control for fertility restoration. Bulked segregant analysis with 215 RAPD primers helped in the identification of putative primers associated with fertility restoration. Co-segregation analysis of eight such primers with Rfl gene revealed two markers, OPK 15700 and OPZ 061300, which flank the Rfl locus on either side at a distance of 8.2 and 2.5 cM, respectively. These DNA markers will be useful in marker-assisted selection for improving the commercial potential of this newly developed CMS-fertility-restorer system for hybrid seed production programs in rapeseed.Key words: oilseed rape, hybrids, cytoplasmic male sterility, fertility restoration, RAPD mapping.

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High-generation near-isogenic lines combined with multi-omics to study the mechanism of polima cytoplasmic male sterility

BMC Plant Biology ◽

10.1186/s12870-021-02852-7 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Benqi Wang ◽

Zunaira Farooq ◽

Lei Chu ◽

Jie Liu ◽

Huadong Wang ◽

...

Keyword(s):

Cytoplasmic Male Sterility ◽

Male Sterility ◽

Higher Plants ◽

Fertility Restoration ◽

Anther Development ◽

Restorer Gene ◽

Joint Analysis ◽

Oil Seed ◽

Oil Seed Rape ◽

Sterility Gene

Abstract Background Cytoplasmic male sterility (CMS), which naturally exists in higher plants, is a useful mechanism for analyzing nuclear and mitochondrial genome functions and identifying the role of mitochondrial genes in the plant growth and development. Polima (pol) CMS is the most universally valued male sterility type in oil-seed rape. Previous studies have described the pol CMS restorer gene Rfp and the sterility-inducing gene orf224 in oil-seed rape, located in mitochondria. However, the mechanism of fertility restoration and infertility remains unknown. Moreover, it is still unknown how the fecundity restorer gene interferes with the sterility gene, provokes the sterility gene to lose its function, and leads to fertility restoration. Result In this study, we used multi-omics joint analysis to discover candidate genes that interact with the sterility gene orf224 and the restorer gene Rfp of pol CMS to provide theoretical support for the occurrence and restoration mechanisms of sterility. Via multi-omics analysis, we screened 24 differential genes encoding proteins related to RNA editing, respiratory electron transport chain, anther development, energy transport, tapetum development, and oxidative phosphorylation. Using a yeast two-hybrid assay, we obtained a total of seven Rfp interaction proteins, with orf224 protein covering five interaction proteins. Conclusions We propose that Rfp and its interacting protein cleave the transcript of atp6/orf224, causing the infertility gene to lose its function and restore fertility. When Rfp is not cleaved, orf224 poisons the tapetum cells and anther development-related proteins, resulting in pol CMS mitochondrial dysfunction and male infertility. The data from the joint analysis of multiple omics provided information on pol CMS’s potential molecular mechanism and will help breed B. napus hybrids.

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iTRAQ-Based Proteomic Analysis of Ogura-CMS Cabbage and Its Maintainer Line

International Journal of Molecular Sciences ◽

10.3390/ijms19103180 ◽

2018 ◽

Vol 19 (10) ◽

pp. 3180 ◽

Cited By ~ 2

Author(s):

Fengqing Han ◽

Xiaoli Zhang ◽

Limei Yang ◽

Mu Zhuang ◽

Yangyong Zhang ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Male Sterility ◽

Molecular Mechanisms ◽

Transcriptome Profiling ◽

Pcr Analysis ◽

Hybrid Seed Production ◽

Protein Species ◽

Absolute Quantitation ◽

Maintainer Line ◽

Cms Line

Ogura cytoplasmic male sterility (CMS) contributes considerably to hybrid seed production in Brassica crops. To detect the key protein species and pathways involved in Ogura-CMS, we analysed the proteome of the cabbage Ogura-CMS line CMS01-20 and its corresponding maintainer line F01-20 using the isobaric tags for the relative and absolute quantitation (iTRAQ) approach. In total, 162 differential abundance protein species (DAPs) were identified between the two lines, of which 92 were down-accumulated and 70 were up-accumulated in CMS01-20. For energy metabolism in the mitochondrion, eight DAPs involved in oxidative phosphorylation were down-accumulated in CMS01-20, whereas in the tricarboxylic acid (TCA) cycle, five DAPs were up-accumulated, which may compensate for the decreased respiration capacity and may be associated with the elevated O2 consumption rate in Ogura-CMS plants. Other key protein species and pathways involved in pollen wall assembly and programmed cell death (PCD) were also identified as being male-sterility related. Transcriptome profiling revealed 3247 differentially expressed genes between the CMS line and the fertile line. In a conjoint analysis of the proteome and transcriptome data, 30 and 9 protein species/genes showed the same and opposite accumulation patterns, respectively. Nine noteworthy genes involved in sporopollenin synthesis, callose wall degeneration, and oxidative phosphorylation were presumably associated with the processes leading to male sterility, and their expression levels were validated by qRT-PCR analysis. This study will improve our understanding of the protein species involved in pollen development and the molecular mechanisms underlying Ogura-CMS.

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Disruption of the bHLH transcription factor Abnormal Tapetum 1 causes male sterility in watermelon

Horticulture Research ◽

10.1038/s41438-021-00695-9 ◽

2021 ◽

Vol 8 (1) ◽

Author(s):

Ruimin Zhang ◽

Jingjing Chang ◽

Jiayue Li ◽

Guangpu Lan ◽

Changqing Xuan ◽

...

Keyword(s):

Transcription Factor ◽

Male Sterility ◽

Molecular Mechanisms ◽

Citrullus Lanatus ◽

Anther Development ◽

Dose Effect ◽

Luciferase Reporter ◽

Bhlh Transcription Factor ◽

Transcriptional Level ◽

Hybrid Seed Production

AbstractAlthough male sterility has been identified as a useful trait for hybrid vigor utilization and hybrid seed production, its underlying molecular mechanisms in Cucurbitaceae species are still largely unclear. Here, a spontaneous male-sterile watermelon mutant, Se18, was reported to have abnormal tapetum development, which resulted in completely aborted pollen grains. Map-based cloning demonstrated that the causal gene Citrullus lanatus Abnormal Tapetum 1 (ClATM1) encodes a basic helix-loop-helix (bHLH) transcription factor with a 10-bp deletion and produces a truncated protein without the bHLH interaction and functional (BIF) domain in Se18 plants. qRT–PCR and RNA in situ hybridization showed that ClATM1 is specifically expressed in the tapetum layer and in microsporocytes during stages 6–8a of anther development. The genetic function of ClATM1 in regulating anther development was verified by CRISPR/Cas9-mediated mutagenesis. Moreover, ClATM1 was significantly downregulated in the Se18 mutant, displaying a clear dose effect at the transcriptional level. Subsequent dual-luciferase reporter, β-glucuronidase (GUS) activity, and yeast one-hybrid assays indicated that ClATM1 could activate its own transcriptional expression through promoter binding. Collectively, ClATM1 is the first male sterility gene cloned from watermelon, and its self-regulatory activity provides new insights into the molecular mechanism underlying anther development in plants.

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RNA-sequencing and bioinformatics analysis of long noncoding RNAs and mRNAs in the asthenozoospermia

Bioscience Reports ◽

10.1042/bsr20194041 ◽

2020 ◽

Vol 40 (7) ◽

Cited By ~ 1

Author(s):

Hui Lu ◽

Dongchuan Xu ◽

Ping Wang ◽

Wenye Sun ◽

Xinhuai Xue ◽

...

Keyword(s):

Rna Sequencing ◽

Regulatory Network ◽

Molecular Mechanisms ◽

Noncoding Rnas ◽

Correlation Coefficients ◽

Long Noncoding Rnas ◽

Pcr Analysis ◽

Mirna Regulation ◽

Qrt Pcr ◽

Positive Correlations

Abstract Asthenozoospermia is one of the major causes of human male infertility. Long noncoding RNAs (lncRNAs) play critical roles in the spermatogenesis processes. The present study aims to investigate the intricate regulatory network associated with asthenozoospermia. The lncRNAs expression profile was analyzed in the asthenozoospermia seminal plasma exosomes by RNA-sequencing, and the functions of differentially expressed genes (DEGs) were analyzed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and DO (Disease Ontology) enrichment analyses. Pearson’s correlation test was utilized to calculate the correlation coefficients between lncRNA and mRNAs. Moreover, the lncRNA–miRNA–mRNA co-expression network was constructed with bioinformatics. From the co-expression analyses, we identified the cis regulated correlation pairs lncRNA–mRNA. To confirm sequencing results with five of the identified DElncRNAs were verified with quantitative reverse-transcription polymerase chain reaction (qRT-PCR). We identified 4228 significantly DEGs, 995 known DElncRNAs, 2338 DEmRNAs and 11,706 novel DElncRNAs between asthenozoospermia and normal group. GO and KEGG analyses showed that the DEGs were mainly associated with metabolism, transcription, ribosome and channel activity. We found 254,981 positive correlations lncRNA–mRNA pairs through correlation analysis. The detailed lncRNA–miRNA–mRNA regulatory network included 11 lncRNAs, 35 miRNAs and 59 mRNAs. From the co-expression analyses, we identified 7 cis-regulated correlation pairs lncRNA–mRNA. Additionally, the qRT-PCR analysis confirmed our sequencing results. Our study constructed the lncRNA–mRNA–miRNA regulation networks in asthenozoospermia. Therefore, the study findings provide a set of pivotal lncRNAs for future investigation into the molecular mechanisms of asthenozoospermia.

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Pentatricopeptide-repeat family protein RF6 functions with hexokinase 6 to rescue rice cytoplasmic male sterility

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1511748112 ◽

2015 ◽

Vol 112 (48) ◽

pp. 14984-14989 ◽

Cited By ~ 60

Author(s):

Wenchao Huang ◽

Changchun Yu ◽

Jun Hu ◽

Lili Wang ◽

Zhiwu Dan ◽

...

Keyword(s):

Cytoplasmic Male Sterility ◽

Male Sterility ◽

Pollen Sterility ◽

Mitochondrial Rna ◽

Hybrid Seed Production ◽

Pentatricopeptide Repeat ◽

Transcript Accumulation ◽

Aberrant Transcript ◽

Family Protein ◽

Restorer Of Fertility

Cytoplasmic male sterility (CMS) has been extensively used for hybrid seed production in many major crops. Honglian CMS (HL-CMS) is one of the three major types of CMS in rice and has contributed greatly to food security worldwide. The HL-CMS trait is associated with an aberrant chimeric mitochondrial transcript, atp6-orfH79, which causes pollen sterility and can be rescued by two nonallelic restorer-of-fertility (Rf) genes, Rf5 or Rf6. Here, we report the identification of Rf6, which encodes a novel pentatricopeptide repeat (PPR) family protein with a characteristic duplication of PPR motifs 3–5. RF6 is targeted to mitochondria, where it physically associates with hexokinase 6 (OsHXK6) and promotes the processing of the aberrant CMS-associated transcript atp6-orfH79 at nucleotide 1238, which ensures normal pollen development and restores fertility. The duplicated motif 3 of RF6 is essential for RF6-OsHXK6 interactions, processing of the aberrant transcript, and restoration of fertility. Furthermore, reductions in the level of OsHXK6 result in atp6-orfH79 transcript accumulation and male sterility. Together these results reveal a novel mechanism for CMS restoration by which RF6 functions with OsHXK6 to restore HL-CMS fertility. The present study also provides insight into the function of hexokinase 6 in regulating mitochondrial RNA metabolism and may facilitate further exploitation of heterosis in rice.

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The Dynamics of Gynodioecy in Plantago lanceolatu L. II. Mode of Action and Frequencies of Restorer Alleles

Genetics ◽

10.1093/genetics/147.3.1317 ◽

1997 ◽

Vol 147 (3) ◽

pp. 1317-1328

Author(s):

Anita A de Haan ◽

Hans P Koelewijn ◽

Maria P J Hundscheid ◽

Jos M M Van Damme

Keyword(s):

Cytoplasmic Male Sterility ◽

Male Sterility ◽

Allele Frequency ◽

Mode Of Action ◽

Male Fertility ◽

Fertility Restoration ◽

Plantago Lanceolata ◽

Allele Frequencies ◽

Male Sterile ◽

Male Fertility Restoration

Male fertility in Plantago lanceolata is controlled by the interaction of cytoplasmic and nuclear genes. Different cytoplasmic male sterility (CMS) types can be either male sterile or hermaphrodite, depending on the presence of nuclear restorer alleles. In three CMS types of P. lanceolata (CMSI, CMSIIa, and CMSIIb) the number of loci involved in male fertility restoration was determined. In each CMS type, male fertility was restored by multiple genes with either dominant or recessive action and capable either of restoring male fertility independently or in interaction with each other (epistasis). Restorer allele frequencies for CMSI, CMSIIa and CMSIIb were determined by crossing hermaphrodites with “standard” male steriles. Segregation of male steriles vs. non-male steriles was used to estimate overall restorer allele frequency. The frequency of restorer alleles was different for the CMS types: restorer alleles for CMSI were less frequent than for CMSIIa and CMSIIb. On the basis of the frequencies of male steriles and the CMS types an “expected” restorer allele frequency could be calculated. The correlation between estimated and expected restorer allele frequency was significant.

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